Process of producing ethyl acetate, ethyl alcohol and other products by fermentation



Patented July 12, 1932 UNITED STATES" PATENT OFFICE IZSAK, OF CENTRAL PARK, NEW JERSEY, AND FRANK A- MCDERMOTT, OF

CLAYMONT, DELAWARE, ASSIGNORS, BY MESNE ASSIGNMENTS, TO E. I. DU IONT DE N EMOURS AND COMPANY, OF WILMINGTON, DELAWARE, A CORPORATION OF DELA- WARE rnocns's or rnonnonve Ernizr. ACETATE, E'IHYL ALCOHOL AND ornnn rnonncrs .BY rnnmnnrarron Ho Drawing. Application filed November This invention relates to the production of ethyl acetate, ethyl alcohol and other products including certain higher alcohols by fermentation.

. It is a well known fact that many species of so-called wild yeasts, especially those of the film-forming type, aside from ordinary ethyl alcohol, also produce fruity odors during fermentation, which by the faculty of 19 smell suggest esters of the lowerfatty acids and aliphatic alcohols. Typical of such wild yeasts are the species designated as Wille'a ammala, M ycoderma m'ni, Myooderma Zactz's,

etc. These are frequently grouped finder the general description of film or surface,

Kahm and ester-yeasts.

To those skilled in the art it is known that all the above described film "yeasts have but relatively meager growth with a subsequent slow fermentation in any concentration of sugar solution, whether it be saccharified grain or molasses as the source.

An object of this invention is the production of ethyl acetate, ethyl alcohol, and other products from fernientable sugars. A further object of our invention is to provide a process whereby a much greater yield of fusel oil and glycerol is obtained. Other objects will appear hereinafter.

We have found by laboratory and plant tests that by treating blackstrap molasses, or beet molasses, or saccharified grain mash, with distillery refuse slop, lively fermentation will be set up immediately upon inoculation with a pure cultureof any of the herein mentioned yeasts. The reason for this perhaps lies in the fact that the distillery slop furnishes enough buffering materials .such as amino-acids, lactic-acid, acetic acid, etc., without which there is no such active fermentation. The amino-acids serve as yeast foods, with the simultaneous production of fusel oil.

It is a well known fact in fermeiitology that the action of all microbes-is a specific one. This notion of specificity has amost im portant consequence in practice as well as in theory. It shows the necessity of employing one and a single organism for the use of a definite end. The Sacohqromycescereoisiw s, 1927. Serial No. 231,990.

type yeasts are not able to utilize as sources of nitrogen, products that are not difl'usible i. e., which cannot pass through the cell wall into the interior of the cellsas its protecclastic enzymes are endo-enzymes, and are not excreted into the medium in which the yeast is growing. For this reason, the production of fusel oil, during their fermentation cannot exceed that theoretically derivable from the amino-acids and lower poly peptides already present in the mash. Other types of yeasts, however, are known which excrete their proteoclastic enzymes into the medium in which they are growing, and hence cannot be conveniently cultured on gelatin media owing to the fact that they digest and liquefy this medium. Among these yeasts is that known as Wz'ZZz'a anomala. This type of film yeast is frequently met with as contaminations in fermentations not conducted under pure culture methods. This micro-organism is a long-celled yeast, which forms a dry, white, wrinkled film on the surface of the sugar solutions, and other appro priate media, produces an active fermentation in its pure culture, accompanied by a fruity odor suggesting ethyl acetate. If the fermented liquid is allowed to stand long enough with access of air, it may completely oxidize the products of fermentation, leaving the solution flat and musty. It liquefies gelatin media slowly, but does not attack good results in acid medium such as a solution of blackstrap molasses or beet molasses in almost any dilution with water, the limiting factor being only that with a very concentrated solution the fermentation is retarded, while with a more dilute solution the fermentation is accelerated, the optimum range being somewhere from 15 to 25 Brix 9 move CO as fast as released( re sulting in a solution. We have further found that by an addition of from 10% to 50% of distillery waste slop, the fermentation isconsiderably accelerated thereby producing much higher yields. We have further found that by reason of the fact that ethyl acetate is very volatile at atmospheric pressures and tempera- 2,000 grams of cane molasses containing about 50%-of total sugars after inversion with acid which reduce Fehlings solution and calculated as invert sugar, are dissolved in 10 litersof water, divided into two equal portions, and sterilized at 5 to 8 lbs. steam pressure in appropriate containers. Each is then inoculated with about 100 c. c. of an active'culture of Wz'llz'a an omala in a portion of thesame solution. One half of the mixture (A) is allowed to ferment without further modification while the other half (B) is aerated by a slow current of airabout 5 bubbles per secondin]'ected through a tube at bottom of the container. (A) will require 7 or 8 days to complete fermentation, while (B) will be completely fermented in about 4 days. On analysis of the two'mashes at the end of fermentation, the following products will be found in approximately the yields indicated, the figures being percents of the total reducing sugar (as invert) initially present in the mashes:

- (A) Non-aerated (B) Aerated Ethyl alcohol 88. to 40. 35'. to 36. Ethyl acetate- 2. to 4. None Fusel oil 0.6 0.8 1 Glycerol 6. to 7. Not determined Acetic acid 3. Not determined Residual reducing substances calculated as invert sugar..- 8. 0 7. 5

Acetaldehyde None observed Trace It is obvious from the above figures that for the production of ethyl acetate, air

should not be passed through the fermenting medium.

According to the end products desired, or for other good reasons the fermentation may be modified in various ways; for instance, (1) a slow air current may be directed over the surface of the fermenting mash, to remore rapid fermentation 2) ammonium salts, phosphates or sulphates, or still other stimulants may be added to hasten the growth of the yeast cells: (3) progressive neutralization of the acidity produced may be secured by suspending, e. g. calcium carbonate in the mash, resulting man increased production of organic acids; (4) the initial mash maybe made up at a very low sugar concentration, and fresh, sterile concentrated mash added either in small portions at intervals or in a continuous stream.

T'Ve have also found that by acidification of the beer, for example, with sulfuric acid, before distillation, an apparent increase in the yield of the ester may be obtained.

As many apparently widely different em bodiments of this invention may be made without departing from the spirit thereof, it is to be understood that we do not intend to be limited by our invention to the specific embodiments thereof except as'indicated in the appended claims.

We claim:

1. The process of producing ethyl acetate and ethyl alcohol by fermentation with yeast fungi thatexcrete their proteoclastic enzymes into an acid fermentable sugar medium which comprises inoculating the medium with fungi of the type of Willia anomala, Myaoderma aim, and Myaoderma lactic controlling the optimum conditions and causing the fermentation of the medium.

2. The process of producinug ethyl acetate and ethyl alcohol with yeast that excrete their proteoclastic enzymes mto a.

sterilized acid fermentable sugar medium which comprises inoculating the medlum with fungi of the type of Willz'a anomala,

Mycodermw' aim", and Myaoderma lactic,

\ prises causing the fermentation of a sterilized acid fermentable sugar medium with the addition of distillery slop, with the herein-described wild yeasts of the type of Willz'a anomala, Mycodema Mai, and Mycoderma lactic. p k

5. The process of producing ethyl acetate and ethyl alcohol, which comprises causing the fermentation of a sterilized acid fermentable sugar medium with the ad 'tion of distillery 'slo with the herein-described wild yeasts, o the type of Willia anomala, lll l coderma vial, and Mycoderma lactic, and acidifying the medium to increase the yield ofesters.

6. In the process of producing ethyl acetate and ethyl alcohol the steps which comprise sterilizing a fermentable sugar medium, adding distillery slop to the medium, and inoculating the medium with the hereindescribed wild yeasts of a type of Willz'a anomala, Mycoderma mm, and Mycodema Zach's. v

7 In the process of producing ethyl acetate and ethyl alcohol the steps which comprise sterilizing a fermentable sugar medium, acidifying the medium, adding to the medium distille slop, and inoculatin the medium with t e herein-described wild yeasts of a type of Wz'llz'a anomala, Mycoderma m'ni, and Mycoderma Zactz's.

In testimony whereof we afiix our signatures.

ALEXANDER IZSAK.

FRANK A. McDERMOTT. 7 

